A biochemical-genetic approach to plasmid DNA replication will be taken. The number of chromosomal and plasmid mutations that deleteriously affect plasmid maintenance will be identified and mapped. Their effect on plasmid replication and segregation will be examined by biochemical techniques. Relaxed mutants of stringently controlled plasmids have been isolated and found to have increased levels of plasmid DNA per chromosome. The biochemical genetic approach will also be used to identify the number and nature of host and chromosomal loci involved in the altered control of plasmid replication. These mutants will also be examined for altered compatibility functions and the specificity of the mutation regarding altered control of other plasmids. The relative effects of specific metabolic inhibitors on plasmid and chromosome replication will also be examined to understand the relationships between macromolecular synthesis and plasmid DNA replication. A physicochemical approach to understanding the unit of segregation involved in plasmid distribution at cell division will be undertaken. The approach consists of examining the "folded" genome of Escherichia coli for the presence of plasmid DNA with subsequent dissociation of plasmid and chromosomal DNA by enzymic means.